Process for producing L-amino acids by fermentation employing a microorganism with resistance to a dipeptide

ABSTRACT

Culturing an L-amino acid producing microorganism belonging to the genus Brevibacterium or corynebacterium and having a resistance to a dipeptide containing glutamic acid or aspartic acid gives L-amino acids in high yield.

This application is a continuation of application Ser. No. 07/464,385, filed on Jan. 12, 1990, now abandoned.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to a process for producing L-amino acids by fermentation and microorganisms for producing L-amino acids.

2. Discussion of the Background

L-amino acids have been widely used as seasonings, medical drugs, feed additives, chemicals, reagents and the like. L-amino acids, which are produced by fermentation on an industrial scale, include L-glutamic acid, L-lysine, L-glutamine, L-arginine, L-phenylalanine, L-alanine, L-threonine, L-isoleucine, L-histidine, L-proline, L-valine, L-serine, L-ornithine, L-citrulline, L-tyrosine, L-tryptophan and L-leucine, etc. As microorganisms utilized for the production of L-amino acids, there are those belonging to the genus Brevibacterium, the genus Corynebacterium, the genus Bacillus, the genus Escherichia, the genus Seratia, the genus Providencia, and the genus Arthorbacter, etc.

It is important to produce L-amino acids at low costs on an industrial scale by enhancing the fermentation yield and accumulation of L-amino acids. To produce L-amino acids industrially at low costs utilizing these various microorganisms, improved breeding of microorganisms has often been used. That is, the L-amino acid production of wild strains per se is extremely poor in many instances, and therefore, methods for imparting nutrient auxotrophy, imparting analog resistance or imparting nutrient auxotrophy in combination with analog resistance, through artificial mutation; or potentiating a gene for amino acid biosynthesis, etc. by genetic recombination, and the like are used to increase the L-amino acid productivity of the wild strain. However, fermentation with conventional strains does not produce L-amino acids in a sufficiently high yield.

Thus, there remains a need for a process which will produce L-amino acids by fermentation in high yield. There also remains a need for microorganisms which produce L-amino acids in high yield.

SUMMARY OF THE INVENTION

Accordingly, it is an object of the present invention to provide a process for producing L-amino acids in high yield by fermentation.

It is another object of the present invention to provide microorganisms which produce L-amino acids in high yield by fermentation.

These and other objects, which will become apparent during the course of the following detailed description, have been achieved by the inventors' discovery that culturing strains having a resistance to a peptide containing glutamic acid or aspartic acid of the formula x-glu, glu-x, x-asp or asp-x, in which x represents an amino acid, produces L-amino acids in improved yields.

That is, one embodiment of the present invention is a process for producing an L-amino acid, which comprises culturing an L-amino acid-producing microorganism belonging to the genus Brevibacterium or the genus Corynebacterium and having a resistance to a peptide containing glutamic acid or aspartic acid in a liquid medium, for a sufficient time to accumulate the L-amino acid in the culture, and collecting the L-amino acid from the culture.

In another embodiment, the present invention relates to L-amino acid-producing microorganisms belonging to the genus Brevibacterium or the genus Corynebacterium and having a resistance to a peptide containing glutamic acid or aspartic acid.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The term L-amino acid as used herein includes L-glutamic acid, L-glutamine, L-lysine, L-arginine, L-phenylalanine, L-threonine, L-isoleucine, L-histidine, L-proline, L-valine, L-serine, L-ornithine, L citrulline L-tyrosine, L-tryptophan and L-leucine, etc. The present invention can even be applied to L-amino acids other than those exemplified herein so long as they are L-amino acids which can be produced by fermentation.

Examples of the peptide of the present invention include tyr-glu, ala-glu, trp-glu, met-glu, gly-glu, glu-gly, glu-leu, glu-his, gly-asp, ala-asp, asp-gly, etc. Strains having a resistance to at least one of these are referred to as peptide-resistant strains in the present invention.

The microorganism belonging to the genus Brevibacterium or the genus Corynebacterium which can be used in the present invention is a variant having the peptide resistance described above and capable of producing an L-amino acid.

To obtain the variants of the present invention, the peptide resistance described above may be induced in the parent strains described below; alternatively, the peptide resistance may also be induced in variants capable of producing an L-amino acid.

Wild strains which can be the parent strains of the variants of the present invention include bacteria belonging to the genus Brevibacterium or the genus Corynebacterium such as a Coryneform producing L-glutamic acid, and are exemplified by the following bacteria.

Brevibacterium flavum ATCC 14067

Brevibacterium lactofermentum ATCC 13869

Brevibacterium divaricatum ATCC 14020

Brevibacterium saccharolyticum ATCC 14066

Corynebacterium glutamicum ATCC 13032

Corynebacterium acetoacidophilum ATCC 13870

For the mutation of these parent strains to the variants of the present invention, a conventional method such as a treatment with N-methyl-N'-nitro-N"-nitrosoguanidine, etc. can be used. Isolation of the variants of the present invention from the mutationtreated bacterial solution can be effected by collecting the strains which can grow in a medium containing the peptide.

Media used for culturing such variants are any conventional media containing carbon sources, nitrogen sources, inorganic ions, substances satisfying nutrient auxotrophy and, if necessary, other organic trace nutrients including vitamins, etc. As carbon sources, there are preferably used carbohydrates such as glucose, sucrose, etc., organic acids such as acetic acid, etc. As nitrogen sources, there are preferably used ammonia water, ammonia gas, ammonium salts, etc. As inorganic ions, potassium ions, sodium ions, magnesium ions, phosphate ions, and the like are appropriately added to the media, as required. Incubation is preferably conducted under aerobic conditions. When the incubation is carried out while adjusting the pH of the medium to a range from 4 to 8, preferably 5 to 7.5, at a temperature of from 25° C. to 37° C., preferably 28° to 34° C., better results can be obtained. Thus, when the present strains are cultured for 1 to 7 days, remarkable amounts of L-amino acids are produced and accumulated in the media. Subsequently using a collecting method using an ion exchange resin, etc., yields crystals of L-amino acid.

Other features of the invention will become apparent in the course of the following descriptions of exemplary embodiments which are given for illustration of the invention and are not intended to be limiting thereof.

EXAMPLES

An example for the mutation of the parent strains to the variants of the present invention and the relationship between the peptide concentration and degree of growth of the present strains are shown below.

Method of mutation

Bacterial cells of Brevibacterium flavum ATCC 14067, which has been grown in a bouillon agar slant at 30° C. for 24 hours, were suspended in M/30 phosphate buffer solution at a cell density of 10⁹ /ml. To the cell suspension was added 200 μg/ml of N-methyl-N'-nitro-N'-nitrosoguanidine. The mixture was maintained at 0° C. for 20 minutes followed by centrifugation. The cells were inoculated on a medium having the composition shown in Table 1 and cultured at 31.5° C. for 2 to 10 days.

                  TABLE 1                                                          ______________________________________                                         Composition of Medium                                                          Component              Content                                                 ______________________________________                                         Glucose                1.0    g/dl                                             Urea                   0.2    g/dl                                             KH.sub.2 PO.sub.4      0.1    g/dl                                             MgSO.sub.4.7H.sub.2 O  0.1    g/dl                                             FeSO.sub.4.7H.sub.2 O  0.002  g/dl                                             MnSO.sub.4.7H.sub.2 O  0.002  g/dl                                             Biotin                 100    μg/l                                          Thiamine hydrochloride 100    μg/l                                          tyr--glu               0.5    g/dl                                             Agar                   2.0    g/dl                                                                    (pH 7.0)                                                ______________________________________                                    

From 20 tyr-glu-resistant strains grown in the agar medium, Brevibacterium flavum AJ 12418 (FERM BP-2205) was obtained and characterized as having high productivity of L-glutamine.

By procedures similar to the mutation above, strains having more improved productivity of amino acids could be obtained using various amino acidproducing strains as the original strain. Representative examples are shown in Table 2.

In addition to the improvement of bacteria capable of producing glutamine, lysine, arginine, glutamic acid, histidine, proline, isoleucine, etc. illustratively shown in Table 2, the present process is also effective for phenylalanine, threonine, valine, ornithine, tryptophan, citrulline, leucine, tyrosine, and serine.

The peptide resistance of the thus-obtained variants was compared with that of the parent strains.

Onto a liquid medium composed of 0.5 g/dl of glucose, 0.2 g/dl of urea, 0.15 g/dl of ammonium sulfate, 0.3 1 g/dl of KH₂ PO₄, 0.1 g/dl of K₂ HPO₄, 0.01 g/dl of MgSO₄.7H₂ O, 0.1 mg/dl of CaCl₂.2H₂ O, 100 μg/l of biotin, 100 μg/l of thiamine hydrochloride, 0.002 g/dl of FeSO₄.7H₂ O, 0.002 g/dl of MnSO₄.7H₂ O and the peptide in the amounts shown in Tables 3-9 and adjusted to a pH of 7.0, there were inoculated suspensions of each of the cells in sterile water, which were obtained by culturing in natural medium (1 g/dl of peptone, 1 g/dl of yeast extract and 0.5 g/dl of NaCl, pH 7.0) in slants for 24 hours. After culturing for 24 hours, the turbidity associated with the growth of bacteria was determined and the degree of growth is expressed in terms of relative growth degree (%) in Tables 3 through 9.

                  TABLE 2                                                          ______________________________________                                                               Peptide  Strain Having                                                         Re-      Improved Yield                                  Amino Acid                                                                              Parent Strain                                                                               sistance Based On Glucose                                ______________________________________                                         Glutamine                                                                               Brevibacterium                                                                              tyr--glu Brevibacterium                                           flavum                flavum AJ 12418                                          ATCC 14067            (FERM BP-2205)                                           Corynebacterium                                                                             ala--glu Corynebacterium                                          acetoacidophilum      acetoacidophilum                                         ATCC 13870            AJ 12419                                                                       (FERM BP-2206)                                  Lysine   Brevibacterium                                                                              val--glu Brevibacterium                                           lactofermentum        lactofermentum                                           AJ 3445               AJ 12420                                                 (FERM P-1944)         (FERM BP-2207)                                           Corynebacterium                                                                             ala--glu Corynebacterium                                          glutamicum            glutamicum                                               AJ 3399               AJ 12421                                                 (FERM P-1615)         (FERM BP-2208)                                  Arginine Brevibacterium                                                                              tyr--glu Brevibacterium                                           flavum                flavum                                                   AJ 3401               AJ 12422                                                 (FERM P-1642)         (FERM BP-2209)                                  Glutamic acid                                                                           Brevibacterium                                                                              tyr--glu Brevibacterium                                           lactofermentum        lactofermentum                                           ATCC 13869            AJ 12423                                                                       (FERM BP-2210)                                           Corynebacterium                                                                             ala--glu Corynebacterium                                          glutamicum            glutamicum                                               ATCC 13032            AJ 12424                                                                       (FERM BP-2211)                                  Histidine                                                                               Brevibacterium                                                                              trp--glu Brevibacterium                                           flavum                flavum                                                   AJ 3620               AJ 12425                                                 (FERM P-2316)         (FERM BP-2212)                                           Corynebacterium                                                                             glu--his Corynebacterium                                          glutamicum            glutamicum                                               AJ 12092              AJ 12426                                                 (FERM P-7273)         (FERM BP-2213)                                  Proline  Brevibacterium                                                                              tyr--glu Brevibacterium                                           flavum                flavum                                                   AJ 11512              AJ 12427                                                 (FERM P-5332)         (FERM BP-2214)                                  Isoleucine                                                                              Brevibacterium                                                                              ala--asp Brevibacterium                                           flavum                flavum                                                   AJ 3686               AJ 12428                                                 (FERM P-2433)         (FERM BP-2215)                                  ______________________________________                                    

                  TABLE 3                                                          ______________________________________                                                 Peptide                                                                Gln       tyr--glu (%)   ala--glu (%)                                          Strain    0      0.05   0.1  0.3 0    0.05 0.1  0.3                            ______________________________________                                         Brevibacterium                                                                           100     85     45   0                                                flavum                                                                         ATCC 14067                                                                     Brevibacterium                                                                           100    100    100  72                                                flavum                                                                         ATCC 12418                                                                     Corynebacter-                    100   70   30   0                             ium                                                                            acetoacido-                                                                    philum                                                                         ATCC 13870                                                                     Corynebacter-                    100  100  100   65                            ium                                                                            acetoacido-                                                                    philum                                                                         AJ 12419                                                                       ______________________________________                                    

                  TABLE 4                                                          ______________________________________                                                 Peptide                                                                Lys*      val--glu (%)   ala--glu (%)                                          Strain    0      0.05   0.1  0.3 0    0.05 0.1  0.3                            ______________________________________                                         Brevibacterium                                                                           100     95     80  10                                                lactofermentum                                                                 AJ 3445                                                                        Brevibacterium                                                                           100    100    100  90                                                lactofermentum                                                                 AJ 12420                                                                       Corynebacter-                    100   50   10   0                             ium                                                                            glutamicum                                                                     AJ 3399                                                                        (FERM P-1615)                                                                  Corynebacter-                    100  100  100  100                            ium                                                                            glutamicum                                                                     AJ 12421                                                                       ______________________________________                                          *When Corynebacterium glutamicum was used, 15 mg/dl of methionine was          supplemented.                                                            

                  TABLE 5                                                          ______________________________________                                                         Peptide                                                        Arg*            tyr--glu (%)                                                   Strain          0      0.05     0.1  0.3                                       ______________________________________                                         Brevibacterium flavum                                                                          100    100       30   0                                        AJ 3401                                                                        Brevibacterium flavum                                                                          100    100      100  95                                        AJ 12422                                                                       ______________________________________                                          *Liquid medium was supplemented with 5 mg/dl of guanine.                 

                  TABLE 6                                                          ______________________________________                                                 Peptide                                                                Glu       tyr--glu (%)    ala--glu (%)                                         Strain    0      0.05   0.1  0.3  0    0.05 0.1  0.3                           ______________________________________                                         Brevibacterium                                                                           100     78    42    0                                                lactofermentum                                                                 ATCC 13869                                                                     Brevibacterium                                                                           100    100    95   80                                                lactofermentum                                                                 AJ 12423                                                                       Corynebacter-                     100   75   40   0                            ium                                                                            glutamicum                                                                     ATCC 13032                                                                     Corynebacter-                     100  100  100  94                            ium                                                                            glutamicum                                                                     AJ 12424                                                                       ______________________________________                                    

                  TABLE 7                                                          ______________________________________                                                 Peptide                                                                His       trp--glu (%)    glu--his (%)                                         Strain    0      0.05   0.1  0.3  0    0.05 0.1  0.3                           ______________________________________                                         Brevibacterium                                                                           100    100     75   36                                               flavum                                                                         AJ 3620                                                                        Brevibacterium                                                                           100    100    100  100                                               flavum                                                                         AJ 12425                                                                       Corynebacter-                     100  100   96  55                            ium                                                                            glutamicum                                                                     AJ 12092                                                                       Corynebacter-                     100  100  100  98                            ium                                                                            glutamicum                                                                     AJ 12426                                                                       ______________________________________                                    

                  TABLE 8                                                          ______________________________________                                                          Peptide                                                       Pro*             tyr--glu (%)                                                  Strain           0      0.05      0.1 0.3                                      ______________________________________                                         Brevibacterium flavum                                                                           100     90       30   0                                       AJ 11512                                                                       Brevibacterium flavum                                                                           100    100       95  60                                       AJ 12427                                                                       ______________________________________                                          *Liquid medium was supplemented with 15 mg/dl of isoleucine.             

                  TABLE 9                                                          ______________________________________                                                          Peptide                                                       Ile              ala--asp (%)                                                  Strain           0      0.05      0.1 0.3                                      ______________________________________                                         Brevibacterium flavum                                                                           100     70       20   0                                       AJ 3686                                                                        Brevibacterium flavum                                                                           100    100       98  70                                       AJ 12428                                                                       ______________________________________                                    

EXAMPLE 1

An aqueous solution medium having a composition of 10% of glucose, 1% of ammonium sulfate, 0.25% of potassium primary phosphate (KH₂ P04), 0.04% of magnesium sulfate, 0.001% of ferrous sulfate, 350 μg/l of thiamine hydrochloride, 5 μg/l of biotin and 0.5 ml/dl of Aji-Eki® which is a soybean protein hydrolysate which accelerates the growth of L-amino acid-producing microorganisms and shortens the required culture time, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in a conventional manner, the various L-glutamine-producing bacterial strains shown in Table 10, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 30 hours at 1200 rpm at an aeration rate of 1/4 volume per minute, while keeping the pH at 6.5 by the addition of ammonia gas. After completion of the fermentation, the yield of L-glutamine produced and accumulated in the solution based on glucose was determined and is shown in Table 10.

                  TABLE 10                                                         ______________________________________                                                                  Yield of L-Glutamine                                  Strain     Property      Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            Wild          29.0                                                  flavum AJ 14067                                                                Brevibacterium                                                                            Imparted with 40.0                                                  flavum AJ 12418                                                                           tyr--glu resistance                                                 Corynebacterium                                                                           Wild          22.5                                                  acetoacidophilum                                                               ATCC 13870                                                                     Corynebacterium                                                                           Imparted with 34.0                                                  acetoacidophilum                                                                          ala--glu resistance                                                 AJ 12419                                                                       ______________________________________                                    

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium flavum AJ 14218 by centrifugation to give a supernatant. From the supernatant, L-glutamine was isolated in a conventional manner using an ion exchange resin to give 19.0 g of L-glutamine as crystals.

EXAMPLE 2

An aqueous solution medium having a composition of 10% of glucose, 2% of ammonium sulfate, 0.1% of potassium primary phosphate, 0.04% of magnesium sulfate, 0.001% of ferrous sulfate, 200 μg/l of thiamine hydrochloride, 500 μg/l of biotin, 5 ml/dl of Aji-Eki®, 1 mg/dl of nicotinamide and 0.1% of DL-alanine, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in a conventional manner, the various L-lysine-producing bacterial strains shown in Table 11, which had been previously grown in bouillon slants at 30° C. for 48 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 7.0 by the addition of ammonia gas. After completion of the fermentation, the yield of L-lysine produced and accumulated in the solution based on glucose was determined and is shown in Table 11.

                  TABLE 11                                                         ______________________________________                                                                  Yield of L-Lysine                                     Strain     Property*     Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            AEC resistance                                                                               16.0                                                  lactofermentum                                                                 AJ 3445                                                                        Brevibacterium                                                                            Imparted with 31.0                                                  lactoermentum                                                                             val--glu resistance                                                 AJ 12420                                                                       Corynebacterium                                                                           met, AEC resistance                                                                          23.0                                                  glutamicum                                                                     AJ 3399                                                                        Corynebacterium                                                                           Imparted with 32.0                                                  glutamicum ala--glu resistance                                                 AJ 12421                                                                       ______________________________________                                          *AEC = S(aminoethyl)-cystein.                                            

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium lactofermentum AJ 12420 by centrifugation to give a supernatant. From the supernatant, L-lysine was isolated in a conventional manner using an ion exchange resin to give 19.2 g of L-lysine as crystals.

EXAMPLE 3

An aqueous solution medium having a composition of 10% of glucose, 4% of ammonium sulfate, 0.1% of potassium primary phosphate, 0.04% of magnesium sulfate, 0.001% of ferrous sulfate, 0.001% of manganese sulfate, 100 μg/l of thiamine hydrochloride, 100 μg/l of biotin, 5 ml/dl of Aji-Eki® and 0.2% of yeast extract, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in a conventional manner, the various L-arginine-producing bacterial strains shown in Table 12, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 7.0 by the addition of ammonia gas. After completion of the fermentation, the yield of L-arginine produced and accumulated in the solution based on glucose was determined and is shown in Table 12.

                  TABLE 12                                                         ______________________________________                                                                  Yield of L-Arginine                                   Strain     Property      Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            gua.sup.-, 2-thiazolyl-                                                                      25.5                                                  flavum AJ 3401                                                                            alanine resistance                                                  (FERM P-1642)                                                                  Brevibacterium                                                                            Imparted with 32.0                                                  flavum AJ 12422                                                                           tyr--glu resistance                                                 (FERM BP-2209)                                                                 ______________________________________                                    

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium flavum AJ 12422 by centrifugation to give a supernatant. From the supernatant, L-arginine was isolated in a conventional manner using an ion exchange resin to give 17.3 g of L-arginine as crystals.

EXAMPLE 4

An aqueous solution medium having a composition of 10% of glucose, 1% of ammonium sulfate, 0.2% of potassium primary phosphate, 0.1% of magnesium sulfate, 0.001% of ferrous sulfate, 0.001% of manganese sulfate, 500 μg/l of thiamine hydrochloride, 5 μg/l of biotin and 1 ml/dl of Aji-Eki®, at pH 7.2, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in an autoclave, the various L-glutamic acid-producing bacterial strains shown in Table 13, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 7.2 by the addition of ammonia gas. After completion of the fermentation, the yield of L-glutamic acid produced and accumulated in the solution based on glucose was determined and is shown in Table 13.

                  TABLE 13                                                         ______________________________________                                                                  Yield of                                                                       L-Glutamine acid                                      Strain     Property      Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            Wild          44.2                                                  lactofermentum                                                                 ATCC 13869                                                                     Brevibacterium                                                                            Imparted with 49.5                                                  lactoermentum                                                                             tyr--glu resistance                                                 AJ 12423                                                                       Corynebacterium                                                                           Wild          40.1                                                  glutamicum                                                                     ATCC 13032                                                                     Corynebacterium                                                                           Imparted with 46.8                                                  glutamicum ala--glu resistance                                                 AJ 12424                                                                       ______________________________________                                    

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium lactofermentum AJ 12423 by centrifugation to give a supernatant. From the supernatant, L-glutamic acid was isolated in a conventional manner using an ion exchange resin to give 35.5 g of L-glutamic acid as crystals.

EXAMPLE 5

An aqueous solution medium having a composition of 10% of glucose, 0.5% of ammonium sulfate, 0.15% of potassium primary phosphate, 0.1% of magnesium sulfate, 0.001% of ferrous sulfate, 0.001% of manganese sulfate, 300 μg/l of thiamine hydrochloride, 350 μg/l of biotin, 5 ml/dl of Aji-Eki® and 0.5% of ammonium acetate, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in an autoclave, the various L-histidine-producing bacterial strains shown in Table 14, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 6.5 by the addition of ammonia gas. After completion of the fermentation, the yield of L-histidine produced and accumulated in the solution based on glucose was determined and is shown in Table 14.

                  TABLE 14                                                         ______________________________________                                                                   Yield of L-Histidine                                 Strain     Property       Based on Glucose (%)                                 ______________________________________                                         Brevibacterium                                                                            2-AT, sulfadiazine,                                                                           7.2                                                  flavum AJ 3620                                                                            cobalamine resistance                                               Brevibacterium                                                                            Imparted with  10.0                                                 flavum AJ 12425                                                                           trp--glu resistance                                                 Corynebacterium                                                                           2-AT resistance                                                                               5.0                                                  glutamicum                                                                     AJ 12426                                                                       Corynebacterium                                                                           Imparted with  9.3                                                  glutamicum glu--his resistance                                                 AJ 12426                                                                       ______________________________________                                    

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Corynebacterium glutamicum AJ 12426 by centrifugation to give a supernatant. From the supernatant, L-histidine was isolated in a conventional manner using an ion exchange resin to give 4.7 g of L-histidine as crystals.

EXAMPLE 6

An aqueous solution medium having a composition of 10% of glucose, 4% of ammonium sulfate, 0.1% of potassium primary phosphate, 0.5% of magnesium sulfate, 0.001% of ferrous sulfate, 0.001% of manganese sulfate, 100 μg/l of thiamine hydrochloride, 350 μg/l of biotin, 1 ml/dl of Aji-Eki® and 35 mg/dl of L-isoleucine, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in a conventional manner, the various L-proline-producing bacterial strains shown in Table 15, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 7.0 by the addition of ammonia gas. After completion of the fermentation, the yield of L-proline produced and accumulated in the solution based on glucose was determined and is shown in Table 15.

                  TABLE 15                                                         ______________________________________                                                                  Yield of L-Proline                                    Strain     Property      Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            ile.sup.-, sulfaguanidine                                                                    21.0                                                  flavum AJ 11512                                                                           resistance                                                          Brevibacterium                                                                            Imparted with 29.0                                                  flavum AJ 12427                                                                           tyr--glu resistance                                                 ______________________________________                                    

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium flavum AJ 12427 by centrifugation to give a supernatant. From the supernatant, 1-proline was isolated in a conventional manner using an ion exchange resin to give 17.5 g of L-proline as crystals.

EXAMPLE 7

An aqueous solution medium having a composition of 10% of glucose, 1% of ammonium sulfate, 0.1% of potassium primary phosphate, 0.04% of magnesium sulfate, 0.001% of ferrous sulfate, 0.001% of manganese sulfate, 100 μg/l of thiamine hydrochloride, 100 μg/l of biotin and 2 ml/dl of Aji-Eki®, at pH 7.0, was charged in an amount of 300 ml into separate small sized glass jar fermenters. After sterilizing in a conventional manner, the various L-isoleucine-producing bacterial strains shown in Table 16, which had been previously grown in bouillon slants at 30° C. for 24 hours, were inoculated thereon. Then, incubation was carried out at 31.5° C. for 48 hours at 1200 rpm at an aeration rate of 1/2 volume per minute, while keeping the pH at 7.3 by the addition of ammonia gas. After completion of the fermentation, the yield of L-isoleucine produced and accumulated in the solution based on glucose was determined and is shown in Table 6.

                  TABLE 16                                                         ______________________________________                                                                  Yield of L-Isoleucine                                 Strain     Property*     Based on Glucose (%)                                  ______________________________________                                         Brevibacterium                                                                            AHV resistance                                                                               8.5                                                   flavum AJ 3686                                                                 Brevibacterium                                                                            Imparted with 13.0                                                  flavum AJ 12428                                                                           ala--asp resistance                                                 ______________________________________                                          *AHV = amino-hydroxy-valeric acid.                                       

The cells were removed from 1 liter of the solution obtained after completion of the fermentation using Brevibacterium flavum AJ 12428 by centrifugation to give a supernatant. From the supernatant, L-isoleucine was isolated in a conventional manner using an ion exchange resin to give 6.5 g of L-isoleucine as crystals.

Thus, as shown by the results given above, according to the present invention, various L-amino acids can be obtained in a good yield.

Obviously, numerous modifications and variations of the present invention are possible in light of the above teachings. It is therefore to be understood that, within the scope of the appended claims, the invention may be practiced otherwise than as specifically described herein. 

What is claimed as new and desired to be secured by Letters Patent of the United States is:
 1. A process for producing an L-amino acid, which comprises culturing an L-amino acid-producing microorganism belonging to a genus selected from Brevibacterium and Corynebacterium and having a resistance to a dipeptide containing glutamic acid or aspartic acid in a liquid medium for a time sufficient to product said L-amino acid, wherein said L-amino acid is selected from the group consisting of L-glutamic acid, L-glutamine, L-lysine, L-arginine, L-isoleucine, L-histidine, and L-proline, and said L-amino acid producing microorganism is Brevibacterium flavum AJ 12418 or Corynebacterium acetoacidophilum AJ 12419 when said L-amino acid is L-glutamine, said L-amino acid producing microorganism is Brevibacterium lactofermentum AJ 12420 or Corynebacterium glutamicum AJ 12421 when said L-amino acid is L-lysine, said L-amino acid producing microorganism is Brevibacterium flavum AJ 12422 when said L-amino acid is L-arginine, said L-amino acid producing microorganism is Brevibacterium lactofermentum AJ 12423 or Corynebacterium glutamicum AJ 12424 when said L-amino acid is L-glutamic acid, said L-amino acid producing microorganism is Brevibacterium flavum ADJ 12425 or Corynebacterium glutamicum AJ 12426 when said L-amino acid is L-histidine, said L-amino acid producing microorganism is Brevibacterium flavum AJ 12427 when said L-amino acid is L-proline, and said L-amino acid producing microorganism is Brevibacterium flavum AJ 12428 when said L-amino acid is L-isoleucine; and recovering the L-amino acid produced.
 2. A culture of a strain of bacteria wherein said strain is selected from the group consisting of Brevibacterium flavum AJ 12418, FERM BP-2205, Corynebacterium acetoacidophilum AJ 12419, FERM BP-2206, Brevibacterium lactofermentum AJ 12420, FERM BP-2207, Corynebacterium glutamicum AJ 12421, FERM BP-2208, Brevibacterium flavum AJ 12422, FERM BP-2209, Brevibacterium lactofermentum AJ 12423, FERM BP-2210, Corynebacterium glutamicum AJ 12424, FERM BP-2211, Brevibacterium flavum, AJ 12425 FERM BP-2212, Corynebacterium glutamicum AJ 12426, FERM BP-2213, Brevibacterium flavum AJ 12427, FERM BP-2214, and Brevibacterium flavum AJ 12428, FERM BP-2215. 